Fig. 1
Discovery and characterization of circFGFR1int2. A Three of the FGFR1-derived circRNAs (hsa_circ_0002352, hsa_circ_0008016, hsa_circ_0005564) recorded in CircBase (n = 17) and two newly identified FGFR1-derived circRNAs by the present study (designated circFGFR1E2/E4/E5 and circFGFR1int2) were expressed in PCa cells. circFGFR1int2 was the most abundantly expressed and most significantly differentially expressed between PCa cells (PC-3, DU145, LNCaP, and 22Rv1) and normal prostate epithelium cell (RWPE-1). B Further characterization of the novel 875-nt circFGFR1int2 by RT-PCR and PCR-sequencing using divergent primers (circ-div-p) spanning splicing junction and convergent primers (circ-con-p). The circFGFR1int2 could be amplified from complimentary DNA (cDNA) obtained by reverse transcription, but not from genomic DNA (gDNA). GAPDH and globin were internal controls for PCR. C circFGFR1int2 was resistant to RNase digestion, as compared to the linear RNA internal control (GAPDH). D Cell fractionation showed that circFGFR1int2 was distributed in both the cytoplasm (Cyto) and the nucleus (Nuc) fractions, whereas FGFR1 mRNA was located in the cytoplasm. ISH (in situ hybridization) showed strong circFGFR1int2 signals (purple) in tumor cell nuclei and cytoplasm of PCa tissues, but only weakly in BPH. E Significant higher circFGFR1int2 (RNA assessed by ISH) and FGFR1 protein (assessed by IHC) levels in PCa (n = 62), but not in BPH (n = 40) tissue samples (P < 0.0001). Violin plots of ISH data were also shown. Bar charts represented semi-quantitative analysis of RT-PCR experiments (n = 3), with. mean ± standard deviation (SD). *P < 0.05, **P < 0.01, ***P < 0.001